Overview

  • Product name

    Anti-WTAP antibody [EPR18744]
    See all WTAP primary antibodies
  • Description

    Rabbit monoclonal [EPR18744] to WTAP
  • Host species

    Rabbit
  • Tested applications

    Suitable for: ICC/IF, IP, Flow Cyt, IHC-P, WBmore details
  • Species reactivity

    Reacts with: Human
    Predicted to work with: Pig, Non human primatesDoes not react with: Mouse
  • Immunogen

    Recombinant fragment within Human WTAP aa 200 to the C-terminus. The exact sequence is proprietary.
    Database link: Q15007

  • Positive control

    • WB: Jurkat, K562, HeLa and HepG2 whole cell lysates; human kidney, thymus and lung lysates. IHC-P:Human endometrium and liver cancer tissues. ICC/IF: Jurkat and HeLa cells. Flow Cyt: Jurkat cells. IP: Jurkat whole cell lysate.
  • General notes

    Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMab® patents.

    This product is a recombinant rabbit monoclonal antibody.

Properties

  • Form

    Liquid
  • Storage instructions

    Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C long term. Avoid freeze / thaw cycle.
  • Storage buffer

    Preservative: 0.01% Sodium azide
    Constituents: PBS, 40% Glycerol, 0.05% BSA
  • Concentration information loading...
  • Purity

    Protein A purified
  • Clonality

    Monoclonal
  • Clone number

    EPR18744
  • Isotype

    IgG
  • Research areas

Applications

Our Abpromise guarantee covers the use of ab195380 in the following tested applications.

The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.

Application Abreviews Notes
ICC/IF 1/500.
IP 1/40.
Flow Cyt 1/600.
IHC-P 1/100. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
WB 1/1000. Detects a band of approximately 50 kDa (predicted molecular weight: 44 kDa).

Milk recommended as blocking agent.

Target

  • Function

    Regulates G2/M cell-cycle transition by binding to the 3' UTR of CCNA2, which enhances its stability. Impairs WT1 DNA-binding ability and inhibits expression of WT1 target genes. May be involved in mRNA splicing regulation.
  • Tissue specificity

    Ubiquitously expressed.
  • Sequence similarities

    Belongs to the fl(2)d family.
  • Post-translational
    modifications

    Phosphorylated upon DNA damage, probably by ATM or ATR.
  • Cellular localization

    Nucleus > nucleolus.
  • Information by UniProt
  • Database links

  • Alternative names

    • DKFZp686F20131 antibody
    • Female-lethal(2)D homolog antibody
    • FL2D_HUMAN antibody
    • hFL(2)D antibody
    • KIAA0105 antibody
    • MGC3925 antibody
    • Mum2 antibody
    • OTTHUMP00000017522 antibody
    • OTTHUMP00000017523 antibody
    • PNAS 132 antibody
    • PNAS-132 antibody
    • PNAS132 antibody
    • Pre mRNA splicing regulator WTAP antibody
    • Pre-mRNA-splicing regulator WTAP antibody
    • Putative pre mRNA splicing regulator female lethal 2D homolog antibody
    • Putative pre mRNA splicing regulator female lethal(2D) antibody
    • Putative pre-mRNA splicing regulator female lethal(2D) homolog antibody
    • putative pre-mRNA splicing regulator female-lethal(2D) antibody
    • Wilms tumor 1 associated protein antibody
    • Wilms tumor 1 associating protein antibody
    • Wilms tumor 1-associating protein antibody
    • Wilms' tumor 1 associating protein antibody
    • Wilms' tumour 1-associating protein antibody
    • WT1 associated protein antibody
    • WT1-associated protein antibody
    • WT1-associating protein antibody
    • wtap antibody
    see all

Images

  • Lane 1: Wild-type HAP1 whole cell lysate (20 µg)
    Lane 2: WTAP knockout HAP1 whole cell lysate (20 µg)
    Lane 3: MOLT-4 whole cell lysate (20 µg)
    Lane 4: K562 whole cell lysate (20 µg)

    Lanes 1 - 4: Merged signal (red and green). Green - ab195380 observed at 50 kDa. Red - loading control, ab18058, observed at 130 kDa.

    ab195380 was shown to specifically react with WTAP in wild-type HAP1 cells as signal was lost in WTAP knockout cells. Wild-type and WTAP knockout samples were subjected to SDS-PAGE. Ab195380 and ab18058 (Mouse anti-Vinculin loading control) were incubated overnight at 4°C at 1/1000 dilution and 1/20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed ab216773 and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed ab216776 secondary antibodies at 1/10000 dilution for 1 hour at room temperature before imaging.

  • All lanes : Anti-WTAP antibody [EPR18744] (ab195380) at 1/1000 dilution

    Lane 1 : Jurkat (Human T cell leukemia cell line from peripheral blood) whole cell lysate
    Lane 2 : K562 (Human chronic myelogenous leukemia cell line from bone marrow ) whole cell lysate
    Lane 3 : HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate
    Lane 4 : HepG2 (Human liver hepatocellular carcinoma cell line) whole cell lysate

    Lysates/proteins at 20 µg per lane.

    Secondary
    All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

    Developed using the ECL technique.

    Predicted band size: 44 kDa
    Observed band size: 50 kDa
    why is the actual band size different from the predicted?


    Exposure time: 3 minutes


    Blocking/Dilution buffer: 5% NFDM/TBST.

    The observed molecular weight is consistent with what has been described in the literature (PMID: 17088532).

  • Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized Jurkat (Human T cell leukemia cell line from peripheral blood) cells labeling WTAP with ab195380 at 1/500 dilution, followed by Goat Anti-Rabbit IgG (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing nuclear staining on Jurkat cell line. The nuclear counter stain is DAPI (blue). 

    Tubulin is detected with Anti-alpha Tubulin mouse MAb (ab7291) at 1/1000 dilution, followed by Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) (ab150120) secondary antibody  at 1/1000 dilution (red).

    The negative controls are as follows:
    -ve control 1: ab195380 at 1/500 dilution, followed by Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) (ab150120) secondary antibody at 1/1000 dilution.
    -ve control 2: Anti-alpha Tubulin mouse MAb (ab7291) at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution.

  • All lanes : Anti-WTAP antibody [EPR18744] (ab195380) at 1/1000 dilution

    Lane 1 : Human kidney lysate
    Lane 2 : Human thymus lysate
    Lane 3 : Human lung lysate

    Lysates/proteins at 10 µg per lane.

    Secondary
    All lanes : Goat Anti-Rabbit IgG Peroxidase Conjugate, specific to the non-reduced form of IgG at 1/100000 dilution

    Developed using the ECL technique.

    Predicted band size: 44 kDa
    Observed band size: 50 kDa why is the actual band size different from the predicted?


    Exposure time: 3 minutes


    Blocking/Dilution buffer: 5% NFDM/TBST.

    The observed molecular weight is consistent with what has been described in the literature (PMID: 17088532).

  • Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HeLa (Human epithelial cell line from cervix adenocarcinoma) cells labeling WTAP with ab195380 at 1/500 dilution, followed by Goat Anti-Rabbit IgG (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing nuclear staining on HeLa cell line. The nuclear counter stain is DAPI (blue). 

    Tubulin is detected with Anti-alpha Tubulin mouse MAb (ab7291) at 1/1000 dilution, followed by Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) (ab150120) secondary antibody at 1/1000 dilution (red).

    The negative controls are as follows:
    -ve control 1: ab195380 at 1/500 dilution, followed by Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) (ab150120) secondary antibody at 1/1000 dilution.
    -ve control 2: Anti-alpha Tubulin mouse MAb (ab7291) at 1/1000 dilution, followed by Goat Anti-Rabbit IgG (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution.

  • Immunohistochemical analysis of paraffin-embedded human endometrium tissue labeling WTAP with ab195380 at 1/100 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution. Nuclear staining on human endometrium is observed. Counter stained with Hematoxylin.

    Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.

    Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

  • Immunohistochemical analysis of paraffin-embedded human liver cancer tissue labeling WTAP with ab195380 at 1/100 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution. Nuclear staining on human liver cancer is observed. Counter stained with Hematoxylin.

    Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.

    Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

  • WTAP was immunoprecipitated from 0.35 mg of Jurkat (Human T cell leukemia cell line from peripheral blood) whole cell lysate with ab195380 at 1/40 dilution. Western blot was performed from the immunoprecipitate using ab195380 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/10000 dilution.

    Lane 1: Jurkat whole cell lysate 10µg (Input).

    Lane 2: ab195380 IP in Jurkat whole cell lysate.

    Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab195380 in Jurkat whole cell lysate.

    Blocking and dilution buffer and concentration: 5% NFDM/TBST.

    Exposure time: 30 seconds.

  • Flow cytometric analysis of 4% paraformaldehyde-fixed Jurkat (Human T cell leukemia cell line from peripheral blood) cells labeling WTAP with ab195380 at 1/600 dilution (red) compared with a rabbit monoclonal IgG isotype control (ab172730; black) and an unlabelled control (cells without incubation with primary antibody and secondary antibody; blue). Goat Anti-Rabbit IgG (Alexa Fluor® 488) at 1/2000 dilution was used as the secondary antibody.

References

This product has been referenced in:

  • Zhu L  et al. Impaired autophagic degradation of lncRNA ARHGAP5-AS1 promotes chemoresistance in gastric cancer. Cell Death Dis 10:383 (2019). WB ; Human . Read more (PubMed: 31097692) »
  • Viphakone N  et al. Co-transcriptional Loading of RNA Export Factors Shapes the Human Transcriptome. Mol Cell 75:310-323.e8 (2019). Read more (PubMed: 31104896) »
See all 6 Publications for this product

Customer reviews and Q&As

1-6 of 6 Abreviews or Q&A

Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample
Pig Tissue sections (Small intestine)
Antigen retrieval step
Heat mediated - Buffer/Enzyme Used: Citric acid
Permeabilization
No
Specification
Small intestine
Blocking step
BSA as blocking agent for 10 minute(s) · Concentration: 2% · Temperature: 21°C
Fixative
Formaldehyde

Mr. Carl Hobbs

Verified customer

Submitted Oct 31 2018

Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample
Common marmoset Tissue sections (Small intestine)
Antigen retrieval step
Heat mediated - Buffer/Enzyme Used: Citric acid
Permeabilization
No
Specification
Small intestine
Blocking step
BSA as blocking agent for 10 minute(s) · Concentration: 2% · Temperature: 21°C
Fixative
Formaldehyde

Mr. Carl Hobbs

Verified customer

Submitted Oct 31 2018

Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample
Human Tissue sections (Testis)
Antigen retrieval step
Heat mediated
Permeabilization
No
Specification
Testis
Blocking step
BSA as blocking agent for 10 minute(s) · Concentration: 2% · Temperature: 21°C
Fixative
Formaldehyde

Mr. Carl Hobbs

Verified customer

Submitted Sep 06 2018

Application
Immunohistochemistry (Frozen sections)
Sample
Baboon Tissue sections (Testis, juvenile (2 years old))
Permeabilization
Yes - 0.1% Triton X-100 in PBS
Specification
Testis, juvenile (2 years old)
Blocking step
BSA as blocking agent for 30 minute(s) · Concentration: 3% · Temperature: 20°C
Fixative
Paraformaldehyde

Mr. Bryan Niedenberger

Verified customer

Submitted Jul 06 2017

Application
Western blot
Sample
Human Cell lysate - whole cell (HeLa)
Gel Running Conditions
Reduced Denaturing (4-15% Bis-Tris)
Loading amount
100 µg
Treatment
siRNA verification
Specification
HeLa
Blocking step
Milk as blocking agent for 30 minute(s) · Concentration: 5% · Temperature: 22°C

Abcam user community

Verified customer

Submitted Nov 29 2016

Application
Western blot
Sample
Human Cell lysate - whole cell (HeLa)
Gel Running Conditions
Reduced Denaturing (4-12%)
Loading amount
10 µg
Specification
HeLa
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C

Abcam user community

Verified customer

Submitted Sep 06 2016

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